Superiority of ABTS over Trinder reagent as chromogen in highly sensitive peroxidase assays for enzyme linked immunoadsorbent assay.

Most of the currently used enzyme immunological assays employ horse radish peroxidase as marker enzyme. A comparison is presented of ABTS (2,2'-azino-di-(3-ethyl benzthiazoline-6-sulphonic acid) and the Trinder reagent as chromogens for the detection of small amounts of solid-phase peroxidase. The formation of chromophore using the Trinder reagent under the conditions described by Gallati (J. Clin. Chem. Clin. Biochem. (1977) 15, 699-703) reaches a plateau after 3-4 h due to H2O2 induced inactivation of the enzyme. In contrast, with suitable temperature and concentrations of H2O2 and ABTS, chromophore production continues in this system for at least 20 h. In an Enzyme Linked Immunoadsorbent Assay for antibodies to myelin basic protein the use of the ABTS/H2O2 substrate system described here gives an assay 14 times more sensitive than the maximum possible with Trinder reagent.